Step-wise assembly, maturation and dynamic behavior of the human CENP-P/O/R/Q/U kinetochore sub-complex.

Kinetochores are multi-protein megadalton assemblies that are required for attachment of microtubules to centromeres and, in turn, the segregation of chromosomes in mitosis. Kinetochore assembly is a cell cycle regulated multi-step process. The initial step occurs during interphase and involves load...

Descripción completa

Guardado en:

Detalles Bibliográficos
Autores principales:	Anja Eskat, Wen Deng, Antje Hofmeister, Sven Rudolphi, Stephan Emmerth, Daniela Hellwig, Tobias Ulbricht, Volker Döring, James M Bancroft, Andrew D McAinsh, M Cristina Cardoso, Patrick Meraldi, Christian Hoischen, Heinrich Leonhardt, Stephan Diekmann
Formato:	article
Lenguaje:	EN
Publicado:	Public Library of Science (PLoS) 2012
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/8ed63ac13a1540678c3a656df8c11a91
Etiquetas:	Agregar Etiqueta Sin Etiquetas, Sea el primero en etiquetar este registro!

id	oai:doaj.org-article:8ed63ac13a1540678c3a656df8c11a91
record_format	dspace
spelling	oai:doaj.org-article:8ed63ac13a1540678c3a656df8c11a912021-11-18T07:05:16ZStep-wise assembly, maturation and dynamic behavior of the human CENP-P/O/R/Q/U kinetochore sub-complex.1932-620310.1371/journal.pone.0044717https://doaj.org/article/8ed63ac13a1540678c3a656df8c11a912012-01-01T00:00:00Zhttps://www.ncbi.nlm.nih.gov/pmc/articles/pmid/23028590/?tool=EBIhttps://doaj.org/toc/1932-6203Kinetochores are multi-protein megadalton assemblies that are required for attachment of microtubules to centromeres and, in turn, the segregation of chromosomes in mitosis. Kinetochore assembly is a cell cycle regulated multi-step process. The initial step occurs during interphase and involves loading of the 15-subunit constitutive centromere associated complex (CCAN), which contains a 5-subunit (CENP-P/O/R/Q/U) sub-complex. Here we show using a fluorescent three-hybrid (F3H) assay and fluorescence resonance energy transfer (FRET) in living mammalian cells that CENP-P/O/R/Q/U subunits exist in a tightly packed arrangement that involves multifold protein-protein interactions. This sub-complex is, however, not pre-assembled in the cytoplasm, but rather assembled on kinetochores through the step-wise recruitment of CENP-O/P heterodimers and the CENP-P, -O, -R, -Q and -U single protein units. SNAP-tag experiments and immuno-staining indicate that these loading events occur during S-phase in a manner similar to the nucleosome binding components of the CCAN, CENP-T/W/N. Furthermore, CENP-P/O/R/Q/U binding to the CCAN is largely mediated through interactions with the CENP-N binding protein CENP-L as well as CENP-K. Once assembled, CENP-P/O/R/Q/U exchanges slowly with the free nucleoplasmic pool indicating a low off-rate for individual CENP-P/O/R/Q/U subunits. Surprisingly, we then find that during late S-phase, following the kinetochore-binding step, both CENP-Q and -U but not -R undergo oligomerization. We propose that CENP-P/O/R/Q/U self-assembles on kinetochores with varying stoichiometry and undergoes a pre-mitotic maturation step that could be important for kinetochores switching into the correct conformation necessary for microtubule-attachment.Anja EskatWen DengAntje HofmeisterSven RudolphiStephan EmmerthDaniela HellwigTobias UlbrichtVolker DöringJames M BancroftAndrew D McAinshM Cristina CardosoPatrick MeraldiChristian HoischenHeinrich LeonhardtStephan DiekmannPublic Library of Science (PLoS)articleMedicineRScienceQENPLoS ONE, Vol 7, Iss 9, p e44717 (2012)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q Anja Eskat Wen Deng Antje Hofmeister Sven Rudolphi Stephan Emmerth Daniela Hellwig Tobias Ulbricht Volker Döring James M Bancroft Andrew D McAinsh M Cristina Cardoso Patrick Meraldi Christian Hoischen Heinrich Leonhardt Stephan Diekmann Step-wise assembly, maturation and dynamic behavior of the human CENP-P/O/R/Q/U kinetochore sub-complex.
description	Kinetochores are multi-protein megadalton assemblies that are required for attachment of microtubules to centromeres and, in turn, the segregation of chromosomes in mitosis. Kinetochore assembly is a cell cycle regulated multi-step process. The initial step occurs during interphase and involves loading of the 15-subunit constitutive centromere associated complex (CCAN), which contains a 5-subunit (CENP-P/O/R/Q/U) sub-complex. Here we show using a fluorescent three-hybrid (F3H) assay and fluorescence resonance energy transfer (FRET) in living mammalian cells that CENP-P/O/R/Q/U subunits exist in a tightly packed arrangement that involves multifold protein-protein interactions. This sub-complex is, however, not pre-assembled in the cytoplasm, but rather assembled on kinetochores through the step-wise recruitment of CENP-O/P heterodimers and the CENP-P, -O, -R, -Q and -U single protein units. SNAP-tag experiments and immuno-staining indicate that these loading events occur during S-phase in a manner similar to the nucleosome binding components of the CCAN, CENP-T/W/N. Furthermore, CENP-P/O/R/Q/U binding to the CCAN is largely mediated through interactions with the CENP-N binding protein CENP-L as well as CENP-K. Once assembled, CENP-P/O/R/Q/U exchanges slowly with the free nucleoplasmic pool indicating a low off-rate for individual CENP-P/O/R/Q/U subunits. Surprisingly, we then find that during late S-phase, following the kinetochore-binding step, both CENP-Q and -U but not -R undergo oligomerization. We propose that CENP-P/O/R/Q/U self-assembles on kinetochores with varying stoichiometry and undergoes a pre-mitotic maturation step that could be important for kinetochores switching into the correct conformation necessary for microtubule-attachment.
format	article
author	Anja Eskat Wen Deng Antje Hofmeister Sven Rudolphi Stephan Emmerth Daniela Hellwig Tobias Ulbricht Volker Döring James M Bancroft Andrew D McAinsh M Cristina Cardoso Patrick Meraldi Christian Hoischen Heinrich Leonhardt Stephan Diekmann
author_facet	Anja Eskat Wen Deng Antje Hofmeister Sven Rudolphi Stephan Emmerth Daniela Hellwig Tobias Ulbricht Volker Döring James M Bancroft Andrew D McAinsh M Cristina Cardoso Patrick Meraldi Christian Hoischen Heinrich Leonhardt Stephan Diekmann
author_sort	Anja Eskat
title	Step-wise assembly, maturation and dynamic behavior of the human CENP-P/O/R/Q/U kinetochore sub-complex.
title_short	Step-wise assembly, maturation and dynamic behavior of the human CENP-P/O/R/Q/U kinetochore sub-complex.
title_full	Step-wise assembly, maturation and dynamic behavior of the human CENP-P/O/R/Q/U kinetochore sub-complex.
title_fullStr	Step-wise assembly, maturation and dynamic behavior of the human CENP-P/O/R/Q/U kinetochore sub-complex.
title_full_unstemmed	Step-wise assembly, maturation and dynamic behavior of the human CENP-P/O/R/Q/U kinetochore sub-complex.
title_sort	step-wise assembly, maturation and dynamic behavior of the human cenp-p/o/r/q/u kinetochore sub-complex.
publisher	Public Library of Science (PLoS)
publishDate	2012
url	https://doaj.org/article/8ed63ac13a1540678c3a656df8c11a91
work_keys_str_mv	AT anjaeskat stepwiseassemblymaturationanddynamicbehaviorofthehumancenpporqukinetochoresubcomplex AT wendeng stepwiseassemblymaturationanddynamicbehaviorofthehumancenpporqukinetochoresubcomplex AT antjehofmeister stepwiseassemblymaturationanddynamicbehaviorofthehumancenpporqukinetochoresubcomplex AT svenrudolphi stepwiseassemblymaturationanddynamicbehaviorofthehumancenpporqukinetochoresubcomplex AT stephanemmerth stepwiseassemblymaturationanddynamicbehaviorofthehumancenpporqukinetochoresubcomplex AT danielahellwig stepwiseassemblymaturationanddynamicbehaviorofthehumancenpporqukinetochoresubcomplex AT tobiasulbricht stepwiseassemblymaturationanddynamicbehaviorofthehumancenpporqukinetochoresubcomplex AT volkerdoring stepwiseassemblymaturationanddynamicbehaviorofthehumancenpporqukinetochoresubcomplex AT jamesmbancroft stepwiseassemblymaturationanddynamicbehaviorofthehumancenpporqukinetochoresubcomplex AT andrewdmcainsh stepwiseassemblymaturationanddynamicbehaviorofthehumancenpporqukinetochoresubcomplex AT mcristinacardoso stepwiseassemblymaturationanddynamicbehaviorofthehumancenpporqukinetochoresubcomplex AT patrickmeraldi stepwiseassemblymaturationanddynamicbehaviorofthehumancenpporqukinetochoresubcomplex AT christianhoischen stepwiseassemblymaturationanddynamicbehaviorofthehumancenpporqukinetochoresubcomplex AT heinrichleonhardt stepwiseassemblymaturationanddynamicbehaviorofthehumancenpporqukinetochoresubcomplex AT stephandiekmann stepwiseassemblymaturationanddynamicbehaviorofthehumancenpporqukinetochoresubcomplex
_version_	1718423992203739136

Step-wise assembly, maturation and dynamic behavior of the human CENP-P/O/R/Q/U kinetochore sub-complex.

Ejemplares similares